Cloning and expression of EBI3 and p28 subunits of human interleukin 27 in E. coli

Interleukin [IL]-27 is a heterodimeric cytokine belonging to IL-12 and IL-23 families, secreted by antigen presenting cells [APCs]. The IL-27 is composed of 2 subunits: Epstein-Barr virus induced gene 3 [EBI3] and p28. IL-27 is an anti-inflammatory cytokine which has an inhibitory effect on Th17 population and suppress the IL-17 expression. It is suggested that IL-27 could be a potent drug candidate for treating auto immune diseases. The EBI3 and p28 subunits of human interleukin 27 were constructed into plasmid vectors; they sub-cloned into pETDuet-1 expression vector in restriction sites of BamHI, SacI and NotI. Subsequently the induction was carried out by 1mM IPTG and the recombinant proteins were confirmed by SDS-PAGE and Western Blot analysis using anti His-tag antibody. The EBI3 and p28 subunits of human interleukin 27were cloned into plasmid vectors. The 28 and 25kDa protein bands were observed on SDS-PAGE and finally confirmed by Western blot technique. In this research p28 and EBI3 proteins were produced successfully

[Diagnosis of microcystin-producing cyanobacteria of Anzali Lagoon using an optimized PCR method]

Production of dangerous cyanotoxins such as microcystin, resulted from cyanobacteria bloom in territorial waters, could cause worldwide issues. We optimized and implemented a PCR method for detection of microcystin-producing cyanobacteria in Anzali Lagoon. Sampling was performed from 20 stations within western, central and eastern parts of the Lagoon. DNA was extracted using DNG-Plus kits. Optimization of PCR test for two universal [23S30R, CYA106F] and two specific primers [mcyA-CdlF, mcyA-CdlR] achieved, and its sensitivity and specificity were evaluated. Moreover, the test was applied for samples from Anzali Lagoon. Optimum PCR yield was obtained at 56°C, and in 0.4 microM concentrations of the primers. Optimized PCR method revealed, which cyanobacteria were present in samples from all of the studied stations of Anzali Lagoon. In addition, microcystin-producing cyanobacteria were observed in 10 out of 20 stations. Anzali Lagoon may have been predominated by dangerous cyanobacteria, and our optimized PCR method can be exerted as an efficient tool for monitoring of microcystin-producing species of cyanobacteria in the lagoons and other water sources